A Redox-active PARP Inhibitor for Lung Transplantation

用于肺移植的氧化还原活性 PARP 抑制剂

• 批准号: 8391286
• 负责人: ANDREW Lurie SALZMAN
• 金额: $ 39.07万
• 依托单位: RADIKAL THERAPEUTICS, INC.
• 依托单位国家: 美国
• 财政年份: 2012
• 资助国家: 美国
• 起止时间: 2012-09-01 至 2014-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8391286
• 关键词: 3-nitrotyrosine; Acute; Acute Lung Injury; Animal Model; Animals; Apoptosis; Area; Breathing; Bronchoalveolar Lavage; Catalysis; Cells; Cellularity; Chairperson; Chemistry; Chlorine; Clinical; Consumption; Cytoprotective Agent; DNA Repair Enzymes; Dose; Drug Delivery Systems; European; Excision; Exhibits; Experimental Models; F2-Isoprostanes; Flushing; Genes; Genetic; Histologic; Histology; Hydrogen Peroxide; IL6 gene; Immunohistochemistry; In Situ; Infarction; Inflammation; Inflammatory; Inhibitory Concentration 50; Injury; Intercellular adhesion molecule 1; Interleukin-1; Interleukin-6; Interruption; Ischemia; Isoprostanes; Left; Left lung; Legal patent; Life; Lipid Peroxidation; Lung; Lung Compliance; Lung Transplantation; MAPK14 gene; MAPK8 gene; Macrophage Inflammatory Protein-1; Measures; Mediating; Messenger RNA; Mitogen-Activated Protein Kinases; Modeling; Morbidity - disease rate; Multiple Organ Failure; Mus; Myocardial Ischemia; Necrosis; Neutrophil Infiltration; Nitrosation; North Carolina; Nuclear; Organ Transplantation; Orphan Drugs; Oxidation-Reduction; P-Selectin; Pathway interactions; Perfadex; Permeability; Pharmaceutical Preparations; Pharmacologic Substance; Pharmacotherapy; Phosphorylation; Pneumonia; Poly Adenosine Diphosphate Ribose; Poly(ADP-ribose) Polymerases; Population; Prevention; Process; Property; Proteins; Pulmonary Edema; Pulmonary Vascular Resistance; Rattus; Relative (related person); Reperfusion Injury; Reperfusion Therapy; Residual state; Respiratory Failure; Resuscitation; Reverse Transcriptase Polymerase Chain Reaction; Risk; Rodent Model; Shock; Shunt Device; Single-Stranded DNA; Small Inducible Cytokine A3; Societies; Sprague-Dawley Rats; Stress; Structure of parenchyma of lung; Sulfhydryl Compounds; TNF gene; TNFRSF5 gene; Technology; Testing; Therapeutic; Thoracotomy; Time; Tissues; Translations; Universities; Vascular Permeabilities; Weight; Zymosan; catalase; catalyst; clinically relevant; commercialization; drug market; exhaust; experience; in vivo Model; indexing; inflammatory marker; inhibitor/antagonist; innovation; lung injury; lung ischemia; medical complication; mimetics; mortality; nitrosative stress; novel; novel therapeutics; oxidant stress; prevent; professor; prophylactic; protein B

DESCRIPTION (provided by applicant): Lung ischemia-reperfusion injury (IRI) is an acute lung injury that contributes to morbidity and mortality following lung transplantation (LTX). IRI i mediated in part by the synthesis of oxidative and nitrosative species that in turn induce DNA single strand breakage and triggering of the nuclear DNA repair enzyme, poly(ADP-ribose) polymerase-1 (PARP-1). Hyperactivation of PARP-1 results in the consumption of its substrate NAD, which in turn exhausts intracellular energetics, leading to ATP depletion and tissue necrosis. PARP-1 activation also induces a widespread expression of pro-inflammatory genes that contribute to vascular permeability, lung edema, neutrophil infiltration, pulmonary shunt, and respiratory failure. In models of IRI injury, genetic deletion or pharmacologic inhibition of PARP-1 potently reduces lung damage, but the extent of protection is subtotal. We propose a more thorough abrogation of IRI-induced tissue damage via the deployment of a bifunctional PARP-1 inhibitor, R-503, formed from the covalent linkage of 2 moieties, each with demonstrated tissue protection: 1) a PARP-1 inhibitor moiety, and 2) a thiol-rich dihydrolipoyl ("DHL") domain that acts as a broad-spectrum redox degradation catalyst. Aim #1: Establish the superiority of the bifunctional PARP-1 inhibitor, R-503, relative to its component domains, in a rat model of warm-ischemic lung IRI. Rat lungs are rendered ischemic in situ for 60 min and reperfused for 4 h. Prior to ischemia, rats are treated with IV R-503, DHL, the monofunctional PARP-1 inhibitor INO-1001, or a combination of DHL and INO-1001. Tissue damage is assessed by histology, levels of PMN infiltration, lipid peroxidation, NF-?B, protein nitrosation, PARP-1 activation, apoptosis, and oxygenation (PaO2). Inflammation is assessed by examining BAL markers of inflammation (protein, PMNs, TNF-¿, and MIP-1¿). Aim #2: Establish the efficacy of R-503 in a syngeneic rat model of orthotopic LTX. Using a translational model of LTX, donor rats are treated IV with R-503 or vehicle 10 min before lung removal. After flushing with cold Perfadex", spiked with R-503 (30 ¿M) or vehicle, donor lungs are stored cold for 12 h before left LTX. Right donor lungs are processed for immunohistochemical analysis of PARP-1 activation. Immediately following LTX with left donor lungs, recipients will receive R-503 or vehicle. The dose of R-503 will be guided by the optimal level elucidated in Aim #1. Recipient rats will be evaluated at 3 different reperfusion times (1, 3, 6 h) for wet/dry weight ratio, graft oxygenation, pulmonary vascular resistance, dynamic compliance, and lung F2¿-isoprostane, histology, and immunohistochemistry for 3-nitrotyrosine and poly(ADP-ribose). Specific analyses will be carried out at 3 time points: at 1 h post reperfusion for I?B¿, phosphorylation of MAP kinases, nuclear p50; at 3 h post reperfusion for RT-PCR to quantify lung mRNA concentrations of TNF-¿, MIP-1¿, and Bcl-2; and at 6 h post reperfusion for determination of BALF cellularity, protein concentration, TNF-¿, MIP-1¿, IL6, and IL1-¿. PUBLIC HEALTH RELEVANCE: Ischemia-reperfusion injury is a major medical complication following lung transplantation and contributes to the high mortality in this population. At present there are no approved prophylactic measures. We are developing a novel drug that targets the basic mechanisms of this condition and will test this agent in two clinically-relevant animal models.

描述（由申请人提供）：肺缺血再灌注损伤（IRI）是一种急性肺损伤，可导致肺移植（LTX）后的发病率和死亡率。IRI i部分由氧化和亚硝化物质的合成介导，氧化和亚硝化物质反过来诱导DNA单链断裂并触发核DNA修复酶聚（ADP-核糖）聚合酶-1（PARP-1）。PARP-1的超活化导致其底物NAD的消耗，这反过来又耗尽细胞内能量，导致ATP消耗和组织坏死。PARP-1激活还诱导促炎基因的广泛表达，促炎基因导致血管通透性、肺水肿、中性粒细胞浸润、肺分流和呼吸衰竭。在IRI损伤模型中，PARP-1的基因缺失或药理学抑制可有效降低肺损伤，但保护程度不完全。我们提出通过部署双功能PARP-1抑制剂R-503来更彻底地消除IR诱导的组织损伤，R-503由2个部分的共价键形成，每个部分具有证明的组织保护作用：1）PARP-1抑制剂部分，和2）富含硫醇的二氢硫辛酰基（“DHL”）结构域，其充当广谱氧化还原降解催化剂。目标一：在大鼠热缺血性肺IRI模型中，确定双功能PARP-1抑制剂R-503相对于其组分结构域的优效性。使大鼠肺原位缺血60分钟并再灌注4小时。在局部缺血之前，用IV R-503、DHL、单功能PARP-1抑制剂INO-1001或DHL和INO-1001的组合处理大鼠。组织损伤评估组织学，中性粒细胞浸润，脂质过氧化反应，NF-？B，蛋白质亚硝化、PARP-1活化、细胞凋亡和氧合（PaO 2）。通过检查炎症的BAL标志物（蛋白质、PMN、TNF-α和MIP-1 β）来评估炎症。目的#2：确定R-503在同系大鼠原位LTX模型中的疗效。使用LTX的转化模型，在肺切除前10分钟用R-503或载体IV处理供体大鼠。在用加入R-503（30 μ M）或溶剂的冷Perfadex冲洗后，将供体肺冷藏12 h，然后进行LTX。处理右供体肺用于PARP-1活化的免疫组织化学分析。在用左供体肺进行LTX后，受体将立即接受R-503或载体。R-503的剂量将由目标1中阐明的最佳水平指导。将在3个不同的再灌注时间（1、3、6 h）评价Wistar大鼠的湿/干重比、移植物 氧合、肺血管阻力、动态顺应性和肺F2 <$-异前列腺素、组织学和3-硝基酪氨酸和聚（ADP-核糖）的免疫组织化学。将在3个时间点进行具体分析：再灌注后1 h I？B，磷酸化 MAP激酶，核p50;在再灌注后3小时进行RT-PCR以定量肺TNF-<$、MIP-1 <$和Bcl-2的mRNA浓度;并在再灌注后6小时测定BALF细胞构成、蛋白浓度、TNF-<$、MIP-1 <$、IL 6和IL 1 <$。 公共卫生关系：缺血-再灌注损伤是肺移植后的主要并发症，并导致该人群的高死亡率。目前还没有批准的预防措施。我们正在开发一种针对这种疾病基本机制的新药，并将在两种临床相关的动物模型中测试该药物。